Lecture-3 Optical Microscopy Introduction Lens formula, image formation and Magnification Resolution and lens defects Basic components and their functions Common modes of analysis Specialized microscopy Techniques Typical examples of applications http://www.youtube.com/watch?v=p2tee17zt4i&list=plkstg-8vpwkzoe4tkVa7f6qmig2hh8meXatn0:46-1:33
Lecture-3 Optical Microscopy • Introduction • Lens formula, Image formation and Magnification • Resolution and lens defects • Basic components and their functions • Common modes of analysis • Specialized Microscopy Techniques • Typical examples of applications http://www.youtube.com/watch?v=P2teE17zT4I&list=PLKstG-8VPWKzOe4TkvA7F6qMlG2HH8meX at~0:46-1:33
Review Problems on Optical Microscopy 1. Compare the focal lengths of two glass converging lenses, one with a larger curvature angle and the other with a smaller curvature angle. 2. List the parameters that affect the resolution of optical microscopes 3. a student finds that some details on the specimen cannot be resolved even after the resolution of the microscope was improved by using the oil immersion objective. The student thinks that the details can be resolved by enlarging a photograph taken with the microscope at maximum magnification Do you agree? Justify your answer. http://www.doitpoms.ac.uk/tlplib/optical-microscopy/questions.php
Review Problems on Optical Microscopy 1. Compare the focal lengths of two glass converging lenses, one with a larger curvature angle and the other with a smaller curvature angle. 2. List the parameters that affect the resolution of optical microscopes. 3. A student finds that some details on the specimen cannot be resolved even after the resolution of the microscope was improved by using the oil immersion objective. The student thinks that the details can be resolved by enlarging a photograph taken with the microscope at maximum magnification. Do you agree? Justify your answer. http://www.doitpoms.ac.uk/tlplib/optical-microscopy/questions.php
http://micro.magnetfsuedu/primer/java/microscopy/immersion/index.html Resolution of a Microscope (lateral) The smallest distance between two specimen points that can still be distinguished as two separate entities dmin =0.61\/NA NA=nsin(a) 入-ⅲ urination wavelength(ight) NA -numerical aperture a-one half of the objective angular aperture n-imaging medium refractive index dmin n 0. 3um for a midspectrum 2 of 0. 55um https://www.youtube.com/watch?v=nzasdncmymoatn5:35-6:00
The smallest distance between two specimen points that can still be distinguished as two separate entities dmin = 0.61l/NA NA=nsin() l – illumination wavelength (light) NA – numerical aperture -one half of the objective angular aperture n-imaging medium refractive index dmin ~ 0.3m for a midspectrum l of 0.55m Resolution of a Microscope (lateral) http://micro.magnet.fsu.edu/primer/java/microscopy/immersion/index.html https://www.youtube.com/watch?v=n2asdncMYMo at~5:35-6:00
Numerical Aperture(NA) NA=1-theoretical Aperture NA=n(sin a) maximum numerical aperture of a lens n: refractive index of the operating with air as imaging medium between the imaging medium the front lens of objective and specimen cover glass Objective lens A Angular aperture Light (≤72 degrees Cone a One half of A-A Specimen cover glass— na of an objective is a measure of its ability to gather light and resolve fine specimen detail at a fixed object distance https://en.wikipedia.org/wiki/angular_aperture http://micro.magnetfsuedu/primer/java/microscopy/immersion/index.html
NA = n(sin ) n: refractive index of the imaging medium between the front lens of objective and specimen cover glass Numerical Aperture (NA) Angular aperture One half of A-A NA=1 - theoretical maximum numerical aperture of a lens operating with air as the imaging medium (72 degrees) https://en.wikipedia.org/wiki/Angular_aperture http://micro.magnet.fsu.edu/primer/java/microscopy/immersion/index.html NA of an objective is a measure of its ability to gather light and resolve fine specimen detail at a fixed object distance. Objective lens Specimen cover glass
Numerical Aperture NA=n(sin a) Oil Immersion and Numerical aperture Imaging Medium Objective Numerical Aperture Comparison Air n=1.0 NA.=0.25 5 NA.=0.75 345 Immersion oil n=1.515 http://ww.youtube.com/watch?v=rskb0j1srnu oil immersion objective use in microscope at0: 33
Numerical Aperture Immersion oil n=1.515 Air n=1.0 http://www.youtube.com/watch?v=RSKB0J1sRnU oil immersion objective use in microscope at~0:33 Imaging Medium NA = n(sin )